Functional Nucleic Acid Nanoparticle-Based ResonanceScattering Spectral Probe

Guiqing Wen & Aihui Liang & Zhiliang Jiang

Received: 5 October 2012 /Accepted: 30 January 2013 /Published online: 9 February 2013# Springer Science+Business Media New York 2013

Abstract Highly sensitive and selective resonance Ray-leigh scattering (RRS) and surface enhanced resonanceRaman scattering (SERRS) spectral detection techniqueare developed by combining the functional nucleic acidincluding aptamer and DNAzyme, and nanoparticle suchas gold/silver (NG/NS) aggregation and catalysis reaction.The recent progress of resonance scattering spectral tech-nologies including RRS and SERRS are reviewed in thispaper.

Keywords Aptamer .DNAzyme .Nanoparticle . ResonanceRayleigh scattering . Surface enhanced resonance Ramanscattering

Introduction

Functional nucleic acid (FNA) mainly includes aptamer andDNAzyme. Aptamers are short single-stranded oligonucleo-tides (20–100 bases), which are selected from synthetic andrandom sequence library in vitro via a combinatorial chem-istry technique known as systematic evolution of ligand byexponential enrichment (SELEX) (Fig. 1). It can bind withhigh affinity and specificity to a wide range of target mol-ecules, such as proteins, polypeptide, organic substance, ormetal ions due to the stable three-dimensional structure ofhairpin [1], pseudoknot [2], stem loop [3], or G-quartet [4](Fig. 2). Such structures result from the shape complemen-tarities recognition between aptamer and target molecules

via van der Waals force, electrostatic interaction, hydrogenbond interaction, shape matching, etc. Aptamers have obvi-ous advantages in analytical and clinical chemistry, includ-ing (1) strong bonding force—their dissociation constantsare only 10−9–10−12; (2) high specificity—the specificity ofaptamer reacting with analyte is higher than that of antibodywith antigen and other analogues did not interfere the reac-tion; (3) easy synthesis in vitro; (4) high stability—they arestable in the process of storage and transit due to less basescontaining in their structure; (5) structural flexibility—theterminal of aptamers can link other functional groups suchas alkyl, base, amidogen, and mercapto group; and (6) thereaction conditions are mild. Therefore, aptamers have beenwidely used in analytical chemistry, clinical diagnosis, anddrug research. DNAzyme are DNA molecules with catalyticactivity. They have outstanding advantages in mimic en-zyme research and heavy metal analysis due to character-istics of easy to be duplicated, synthesized, and modified;high thermal stability; and low hydrolytic rate [5–9]. Thetwo kinds of FNA have been widely used as very importantanalytical reagents in spectral analysis, electroanalysis,chromatographic analysis, and biosensor. Especially, FNAhave rapid development in nanoanalysis in recent years.

Nanoparticles have been widely applied in analyticalchemistry due to the small size effect, quantum effect, andmacroscopic quantum tunneling effect. Nanogold/nanosilver(NG/NS) have presented unique optical and electrical featuresand high compatibility. It has been widely used in nanoelec-tronics, optics, catalysis, and bioanalysis because it can com-bine with many biomolecules such as nucleic acid, protein,and enzyme. On one hand, unmodified NGs/NSs can be usedin resonance scattering (RS) spectral analysis. On the otherhand, antibody (aptamer)-modified NG/NS have better selec-tivity as RS spectral probes. NG has played an important rolein nanoanalysis and become the fourth label following radio-active element, enzyme, and fluorescein [10–12] because it is

G. Wen :A. Liang : Z. Jiang (*)Key Laboratory of Ecology of Rare and Endangered Speciesand Environmental Conservation of Education Ministry, GuangxiKey Laboratory of Environmental Pollution Control Theoryand Technology, College of Environment and Resource,Guangxi Normal University, Guilin 541004, Chinae-mail: zljiang@mailbox.gxnu.edu.cn

Plasmonics (2013) 8:899–911DOI 10.1007/s11468-013-9489-y

easy to be prepared, stable and has high catalytic activity. NSis cheaper compare to NG. It is a highly active and commonlyused substrate for surface enhanced Raman scattering (SERS)spectroscopy. NS has played an important role in SERS andsurface enhanced resonance Raman scattering (SERRS) quan-titative analysis [13, 14], and has also been used in resonance

Rayleigh scattering (RRS). In this article, the background ofthe RRS and SERS analytical techniques, aptamer-based RRSmethod, DNAzyme-based RRS method, aptamer-basedSERRS quantitative analysis, and aptamer-based SERRSquantitative analysis were reviewed [8, 9, 11, 13–19].

Background of the RRS and SERS Analytical Techniques

When the incident light irradiates the samples, there arethree probable ways such as transmission, absorption,and scattering. The wavelength of most of the scatteringlight is the same as that of the incidence light called asRayleigh scattering, while small part has wavelengthshift due to molecular vibration and rotation. The latteris called as Raman scattering light (Fig. 3). The signalsof Rayleigh scattering are strong while those of Ramanscattering are very weak. Rayleigh scattering analysis isa simple, rapid and sensitive technique that has beenutilized in ionorganic and organic analysis, combiningwith various chemical reactions. But Raman scattering

Fig. 1 SELEX technique

hairpin pseudoknot

bulge G-quartet

Fig. 2 Structures of aptamer

900 Plasmonics (2013) 8:899–911

is rarely used because its sensitivity is low. RS techniqueshown in this paper is mainly combining with nanopar-ticles. It mainly includes RRS spectroscopy which isbased on elastic scattering including Rayleigh scatteringand Mie scattering, and SERS spectroscopy which isbased on inelastic scattering. When some Raman activemolecules are attached to the surface of a nanoparticlesuch as nanosilver or nanogold, the Raman scatteringsignal enhanced 106 times that called as SERS. TheSERS has the advantages of high sensitivity, large infor-mation quantity, non-disturbance by water, and multi-component measurement. It shows a good applicationperformance in immune detection and aptamers assay[20–23]. Compare to RRS, SERS has less quantitativeanalysis because rarely active substrates can be used inthis quantitative analysis though many solid substratesfor SERS have been reported.

Resonance Rayleigh Scattering Spectral Analysis

The research and analytical application of RRS is later thanthat of Raman scattering [24–26] though Rayleigh scatteringphenomenon was observed by Rayleigh as early as 1899.RRS closely correlate with the characteristic of nanoparticlesurface, especially with the resonance absorption of surface

plasmon. Thus, it is also called as surface plasmon reso-nance Rayleigh scattering (SPR-RS). The size, shape, andproximity of nanostructures all affect the magnitude of theSPR-RS. By selecting suitable conditions and reducing theeffect of interference on nanoparticle SPR-RS, the selectiv-ity of RRS method has been improved in virtue of highlyselective aptamer reaction and enzymatic reaction. Based onthese applications in FNA, RRS spectral analysis has beendeveloped rapidly.

Aptamer-Based RRS Method

There were reports about the application of NG/NSaptamer probe in spectrophotometry, electrochemicalanalysis, and fluorescence spectroscopy previously. ButNG/NS have been used in detection combining with RRSjust in recent years. Aptamers were used to modify NGto obtain an aptamer-NG probe for K+. When theaptamer-NG interacted with K+, the aptamer reacted withK+ to form stable complex and NGs was released. Inhigh concentration of electrolyte, the released NGs wereaggregated to cause the RRS intensity enhanced. Thus,an aptamer–NGs RRS spectral assay for K+ was pro-posed. Based on this phenomenon, the RRS spectralassay for Hg2+ using aptamer-modified NGs as probewas developed. Furthermore, after the large NGs clusters

Fig. 3 Rayleigh scattering and Raman scattering. Left scattering model; right scattering energy level

Fig. 4 G-quartet structure of nanogold combining with aptamer

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were removed by membrane filtration, the excessaptamer-NG in the filtrate solution exhibits a catalyticeffect on the Cu2O particle reaction between NH2OH andCu(II). The decreased RS intensity of Cu2O particle hada linear response to Hg2+ concentration. So, an aptamer-modified NG catalytic RRS method was built [27]. Themethod of Cu enhancement is cheap, stable, reproducible,

and sensitive and it has the advantage over Ag enhancementor Au enhancement.

Zhang et al. [28] has developed a new RRS method todetermine urinary adenosine (AD). The method is baseon specific recognition for AD aptamer and signal am-plification of APT, which is combining with the labeledNGs (Fig. 4). The RRS spectra indicated that both scat-tering signals of the AD and DNA-NGs systems are veryweak over the scanning wavelength range of 200–700 nm. However, the RRS intensity enhanced largelywith the addition of AD and appeared at three obviousRRS peaks at 450, 470, and 495 nm (Fig. 5). There waslinear relationship between the RRS signal at 495 nmand AD concentration of 6–115 nmol/L, and the detec-tion limit is 1.8 nmol/L. The method was applied todetermine urinary adenosine successfully and the resultsagreed well with those of HPLC. Aptamers-modifiedNG/NS reactions have been combined with nanocatalysisreactions to develop new RRS spectral assays in recentyears. The analytic application of nanoparticle–aptamer-based RRS spectral assays was listed in Table 1.

DNAzyme-Based RRS Method

The RRS, label-free detection of an analyte is probably themost favorable method from the end user perspective. Such

Fig. 5 RRS spectra of the AD detection system. 1, AD; 2, DNA-NGs;3–8, DNA-NGs-AD, the concentration of AD (in nanomoles per liter):0, 25, 40, 55, 70, and 115

Table 1 FNA–nanoparticle RRS spectral assays

Analyte Principle Linear range Detectionlimit

Sample Reference

Thrombin K+ combine with Apt-AuReNP to make G-quartet structure 0.115–6.93 nM 13 pM Human plasma [30]

Melamine AgNP-ssDNA reaction combine with the Cu(II)catalytic reaction

6.31–378.4 μg/L 3.1 μg/L Milk [31]0.02–1.06 μg/L 0.01 μg/L

Adenosine K+ combine with Apt-AuNP to make G-quartet structure 6–115 nM 1.8 nM Urinary [28]

ATP AuNP-dsDNA reaction combine with the Cu(II) catalyticreaction

6.6–110 nM – Synthesissamples

[32]2.2–220 nM 0.5 nM

Hg2+ AuRuNP-ssDNA reaction combine with the AuRuNP catalyticreaction

0.0067–3.3 nM 0.0048 nM Water, tea [33]2.7–133.3 pM 1.5 pM

Hg2+ AuNP-hsDNA-Cu(II)-NH2OH catalytic reaction 0.4–400 nM 0.2 nM Wastewater [34]

Hg2+ AuNP-hsDNA-glucose-Cu(II) catalytic reaction 1–833 nM 0.3 nM Wastewater [35]

Hg2+ AuNP-ssDNA-HAuCl4-AA catalytic reaction 0.00008–0.888 ng/mL

0.000034 ng/mL

Wastewater [36]

Hg2+ AuReNP-ssDNA reaction combine with the Te catalyticreaction

0.067–33. 3 nM 0.04 nM River water [37]0.00133–0.267 nM 0.9 pM

Pb2+ Pb2+ combine with Apt-AuNP to make G-quartet structure 0.07–2 nM 0.03 nM Wastewater [38]

Pb2+ Pb2+ combine with Apt-AuPdNP to make G-quartet structure 0.08–42 nM 0.04 nM Wastewater [39]

K+ K+ combine with Apt-AuNP to make G-quartet structure 0.67–3,350 μM 0.3 μM Human plasma [40]

UO22+ AuPdNP-dsDNA reaction combine with AuPdNP-dsDNA-

NaH2PO2-NiCl2 catalytic reaction40–250 pM 20 pM Water [41]5.0–50 pM 2 pM

UO22+ AuNP-dsDNA-HAuCl4-AA catalytic reaction 3.35–23.45 pM 0.1 pM River water,

lake water[11]

AuNP gold nanoparticle, AgNP silver nanoparticle, ssDNA single-strand DNA, AuPdNP AuPd nanoparticle, AuRuNP AuRu nanoparticle, Aptaptamer, CTMAB cetyltrimethyl ammonium bromide

902 Plasmonics (2013) 8:899–911

a detection method is possible with DNAzymes and NGs.DNAenzyme is the important FNA reagent for the determi-nation of some heavy metals and has bright prospect onRRS analysis. Zhang et al. [11, 29] have developedDNAzyme-based RRS spectroscopy for UO2

2+ (Fig. 6).The hybrid DNA was isolated and cleaved to a single-strand DNA (ssDNA) which was combined with NGs andstabilize it. In the absence of UO2

2+, the NGs were aggre-gated to larger particles in high concentration of electrolyte.The dilution of enzymatic fluid exhibits a catalytic effect onthe gold particle reaction between ascorbic acid andHAuCl4, and the product of gold particles exhibited a RRSpeak at 620 nm as in Fig. 7. With the concentration ofUO2

2+ increasing, the concentration of NGssDNA increasedin the cracking reaction solution, the catalytic product ofgold particles increased, and the RS intensity at 620 nmenhanced linearly. The linear range is 3.35–23.45 pmol/LUO2

2+, and the detection limit is 0.1 pmol/L. The analyticapplication of nanoparticle–DNAzyme-based RRS spectralassays was listed on Table 1.

SERRS Spectral Analysis

Raman spectroscopy is a vibrational technique; it can beused to identify functional group in the molecules andreflect its microstructure characteristics. However, the weaksignals of normal Raman (as 10−10 times of incident light)result in poor sensitivity. The research of Fleischmann [42],Van Duyne [43], and Creighton [44] groups has also shownthat the Raman signals of pyridine that adsorbed on thesurface of roughened silver electrode had been amplified106 times than that of non-surface-enhanced pyridine. TheSERRS is highly sensitive and the signals of H2O are veryweak and cannot interfere with the signals of the analytes.The intensity of normal Raman scattering (NRS) can beshown as I1(ωs)∝N1I1(ωi)σf, where N1 is the moleculesnumber of incident laser irradiated and σf is the Ramanscattering cross section of free molecules (Fig. 8). Thescattering intensity depends linearly on the numbers ofscattering molecules, the intensity of incident light, and theRaman scattering cross section of free molecules. The

Fig. 6 DNAzyme-based RRSspectroscopy for UO2

2+

Plasmonics (2013) 8:899–911 903

scattering cross section of single molecule (about 10−30

cm2) is smaller than that of luminescent dye (coloredgroups) by 14 logarithms (10−16cm2). Since the valueof σf is so small, NRS is weak. For SERR, its intensitycan be shown as I2(ωs)∝N2I2(ωi)I2(ωs)∝N2· I1(ωi)σaF(ωi) ·F(ωs), where N2 is the number of scattering moleculesinvolved in SERS process. F(ωi) and F(ωs) are fieldenhancement factor of laser incident field and Stokesscattering field, respectively, and σa is the enhancedcross section of adsorbed molecules (chemical enhancedcross section). The diameter of metal particles in Fig. 8bis at the range of 10–100 nm. In the system of dyeadsorbed molecules and silver, the number (N2) of par-ticles involved in SERS is far fewer than that (N1) ofmolecules involved in NRS. However, the Raman en-hancement factor of single molecule which is contributedby σaF(ωi)F(ωs) (enhancement cross section of adsorbedmolecule σa−200) is 1014–1015. So, SERS is much moresensitive than NRS [45, 46]. The SERS technique has

been widely used in biology [45–47], medicine [23,48–51], and environment [52, 53] due to high sensitivity,selectivity, and rich information. During the study ofSERS, people have found that resonance Raman scatter-ing appeared when the wavelength of excited light wasequal to that of electronic transition of Raman activemolecules. It is more difficult to get resonance Ramanscattering, though its signal is about 106 times as NRSsignal. Furthermore, the wavelength of excited light mustmatch the absorption of chromophore in analyte mole-cules. The intensity of exciting light and Raman scatter-ing are both related to the thickness of samples and makethe quantitative analysis complicated.

According to SERS theory, if Raman molecule thatadsorbed on aggregated nanoparticles surface has chro-mophore, SERRS appeared when the wavelength ofexciting light was equal to that of absorption. Thesignals are enhanced largely and the high sensitivityhas been gained. In SERS detection, since the SERSmolecules are mainly rhodamine dyes and mercaptopyr-idine that have chromophore, it is SERRS in fact.Therefore, SERRS was introduced in this paper as title.During the SERS development of over 30 years, itsdisadvantages were as notable as the advantages andare as follow: (1) In most present studies, nanosilverparticles were fixed on the solid substrates such as glassand silica to get rough substrates with high SERSactivity. But the reproducibility of these kinds of SERSactive substrates was unsatisfactory, and it is very diffi-cult to get SERS active substrates of good reproducibil-ities. (2) Time–space characteristics of SERS activesubstrates are complicated and the thermal effects oflaser excitation should not be ignored. (3) The SERSenhancement solely depends on the space between theanalytes and substrates and change of coverage ratio ofanalytes on substrates surface [54, 55]. Therefore, as anew molecule scattering spectral technology, the SERSmethods are facing new tasks as well as challenges anddevelop gradually. The SERS quantitative methods withhigh sensitivity and good reproducibility depend on thestable NGs/NSs and it is an important way to modifyNGs/NSs with FNA.

Wavelength (nm)

Inte

nsit

y (a

.u.)

f

e

d

c

b

a

Fig. 7 RRS spectra of the NGssDNA-HAuCl4-AA catalytic system. a,32 μmol/L HAuCl4-3.3 mmol/L HCl-5 mmol/L CTMAB-34 mmol/LAA; b, a-7.74 μg/mL NG-1.07 nmol/L dDNA-3.35 pmol/L UO2

2+; c,a-7.74 μg/mL NG-1.07 nmol/L dDNA-6.7 pmol/L UO2

2+; d, a-7.74 μg/mL NG-1.07 nmol/L dDNA-10.05 pmol/L UO2

2+; e, a-7.74 μg/mL NG-1.07 nmol/L dDNA-20.1 pmol/L UO2

2+; f, a-7.74 μg/mL NG-1.07 nmol/L dDNA-23.45 pmol/L UO2

2+

Fig. 8 Sketch map of normal Raman scattering (a) and SERS (b)

904 Plasmonics (2013) 8:899–911

Fig. 9 SERRS-based aptamers sensor for α-thrombin

Fig. 10 Aptamer biosensor for cocaine

Plasmonics (2013) 8:899–911 905

Aptamer-Based SERRS Analysis

Cho et al. [56] have developed a SERRS-based aptamersensor with high sensitivity, high specificity, and good sta-bility (Fig. 9). It can identify and detect α-thrombin ofhuman with a detection limit of 100 pmol/L. He researchgroup [57] has developed a “two-step” aptamers SERRSmethod to detect ricin in liquor foods. In this method, ricinB chain can be captured by aptamer which are conjugatedpairs of nanocrystalline silver, and the values of SERRSsignals are changed (Fig. 10). This kind of aptamer sensorthat made by conjugation is easy to prepare and high sensi-tive. It can be widely used to detect ricin in all kinds ofliquor foods. With this method, the detection limit of ricin insaline, orange juice, and milk is 10, 50, and 100 ng/mL,respectively. Yu et al. [58] have developed a novel aptamersensor for cocaine. They found that the target molecules

which were surface-affinitive to the SERRS substrate hadSERRS signals after combining with a substrate. Under theselected conditions, the detection limit of cocaine is 1 μmol/L.

Heavy metal pollution has become one of the hotspot. Asa highly sensitive spectral technique, SERRS sensor hasbeen used to detect trace Hg2+. Chung et al. [59] haveprepared a novel SERRS spectral probe for the detectionof 50 nM Hg2+ (Fig. 11). One end of the aptamer moleculethat was modified by tetramethylrhodamine (TAMRA) waslabeled on the shell of NGs to form TAMRA–aptamer–NGscomplex. The complex interacted with Hg2+ to form stablehairpin structure with T-Hg2+-T mismatches, which resultedin the adsorption of another end of aptamer on the shell ofNGs. The higher Hg2+ concentration is, the more TAMRAadsorbed on the shell of NGs are, and the larger the SERRSintensity of TAMRA are. The spectra in Fig. 12 show thatthe most prominent peak of TAMRA is observed at

Fig. 11 Schematic descriptionof the SERRS sensor for Hg2+

ion detection

Fig. 12 SERRS spectraobtained from DNA-modifiedgold microshells at a variety ofHg2+ ion concentrations

906 Plasmonics (2013) 8:899–911

Fig. 13 Schematicrepresentation showing theprotocol for the label freedetection of Hg(II) usingtryptophan-protected popcorn-shaped gold nanoparticles

Table 2 Application of SERS to aptamer analysis

Analyte SERS molecule Substrate Linear range Detection limit Ref.

Adenosine Tetramethyl rhodamine AuNP sol 0.02–2 μmol/L 10 nmol/L [15]

Thrombin 4-Aminothiophenol Au film 0.001–1 mmol/L 1 μmol/L [56]

ATP MGITC Gold nanostar@Raman 0–2.0 nM 12.4 pM [61]

label@SiO2 CSNP

α-Thrombin α-Thrombin aptamer AuNP 100 pmol/L–1 μmol/L 100 pmol/L [62]

Protein AgNP dimers [63]

Cocaine Cocaine aptamer AuNP – – [58]

Cocaine Cocaine aptamer AgNP sol Semiquantitative analysis 1 μmol/L [63]

1–3,200 μmol/L

Pathogen Pathogen aptamer AuNW 10 pmol/L–10 nmol/L 10 pmol/L [64]

α-Thrombin Rhodamine 6G Au film Semiquantitative analysis 0.5 nmol/L [65]

0.5–120 nmol/L

Ricin Ricin aptamer Nanocrystalline silver – 10 ng/mL, saline [57]

– 50 ng/mL, orange juice

– 100 ng/mL, milk

Melamine Aptamer AgNP sol 6.3–403.6 μg/L 1.2 μg/L, milk [66]

DNA TAMRA AgNP sol 0.2–2.0 nM 77 pM [67]

Hg2+ AuNW [68]

Hg2+ TAMRA-label aptamer AgNP sol 5 nmol/L–5 μmol/L 5 nmol/L [69]

Pb2+ Pb2+ aptamer Au film 0.1–10 nmol/L 100 pmol/L [70]

AuNW gold nanowire, AgNP sol silver nanoparticle sol, ATP adenosine triphosphate, MGITC malachite green isothiocyanate, CSNP core–shellnanoparticle, TAMRA tetramethylrhodamine.

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1,650 cm−1, so as the change in SERRS intensity as afunction of Hg2+ ion concentration. The increasing trendof SERRS signals in response to augmented Hg2+ ion con-centration is in good accordance with the expected mecha-nism shown in Fig. 11. This result strongly supports thesuggested scenario that the locked hairpin structure and theTAMRA Raman active probe located close to the gold

microshell surface leads to an increased SERRS signal fromthe TAMRA Raman reporters. Based on the SERRS effect

500 1000 1500 20000

100

200

300

400R

aman

Inte

nsity

(cps

)

Raman Shift(cm-1)

b

a

Fig. 14 SERRS spectra of the UO22+-dsDNA-GN-RhG system. a,

0.76 nmol/L dsDNA-11.8 μg/mL GN-1.83 μmol/L RhG; b,a-100 nmol/L UO2

2+

Fig. 15 The principle of label-free NGs-DNAzyme-SERRSmethod for UO2

2+

Table 3 Comparing of RRS and SERRS

RRS SERRS

Elastic scattering, resonancebetween the excited photonand the surface electron ofnanoparticles

Inelastic scattering, resonancebetween the excited photonand the electron in chromophoreadsorption on the nanoparticles

All reported methods arequantitative analysis,nanoparticles can beutilized as RRS nanoprobe

Most reported methods arequalitative analysis, andquantitative analysis is less,nanoparticles were utilized assubstrate and the chromophorewas used as molecular probe

Measured in solution withgood precision

Measured in solid substrate withbad precision. If the SERRSrecording carried out in solutionits precision would be improved

High sensitivity, selectivitythat should be coupledwith antibody, simplicity

High sensitivity, high selectivitywith rich information,complexity

Middle cost of the equipment,and low cost of the operation

Expensive cost of the equipmentand high cost of the operation,but exception the portableRaman spectroscope

908 Plasmonics (2013) 8:899–911

of tryptophan at 1,467.8 cm−1 and the quenching effect oftryptophan–Hg(II) complex on the NGs, Ray et al. [60] havereported a SERRS sensor for detection of Hg2+ (Fig. 13),with a detection limit of 5 ppb. The application of SERSspectral techniques on aptamer analysis are listed on Table 2[15, 56, 57, 61–67].

DNAzyme-Based SERRS Analysis

Labeling the DNAzyme with Raman tags costs extra timeand materials and they may potentially interfere with theenzymatic activities. Therefore, SERRS sensors withoutthese external tags will have an advantage in these aspects.Recently, several label-free SERRS sensors have been de-veloped by several groups. Base on metal ion catalysis ofDNA cracking, the enzyme reaction was combined with theNG/NS aggregation to develop SERRS methods [68–70].Rhodamine 6G (RhG) is a kind of cationic dye with highSERRS activity that appeared eight strong SERRS peaks atthe Raman shift of 611, 773, 1,185, 1,311, 1,360, 1,510,1,570, and 1,649 cm−1 (Fig. 14). In the conditions of pH 5.5buffer solution (2-(4-morpholino)ethanesulfonic acid) and0.019 mol/L NaCl, NGs cannot be protected by dsDNAand form tight aggregated NGs (TANG). The interactionbetween TANG and RhG was weak and exhibited weaksignals (Fig. 15a). After adding UO2

2+, it can cleave thesubstrate strand of DNA and release a ssDNA. The ssDNAcan be adsorbed on the surface of NGs to form NGs-ssDNAcomplex. Upon the addition of RhG, NGs-ssDNA wasadsorbed to the surface of RhG and exhibited strong SERRSpeak at 1,360 cm−1 (Fig. 15b). The higher the UO2

2+ con-centration, the more ssDNA are produced. And the moreNGs-ssDNA-RhG are, the larger the SERRS intensity is.Thus, the SERRS spectral method for UO2

2+ was developedand the linear range was 5–125 nmol/L.

Conclusion

The RS spectroscopy is a new technique of trace analy-sis. RRS is based on SPR-RS effect of nanoparticles andis the resonance of excited light with the SPR electron. Itis highly sensitive and the selectivity has been improvedby combining with FNA. SERRS appeared when theelectrons in vibrational energy level of colored moleculesthat adsorbed on the surface of nanoparticles resonatedwith the excitation photon. SERRS technology hasadvantages of high sensitivity, good selectivity, and richinformation (Table 3). Its combination with FNA hasimproved the analytical properties such as accuracy andbroadened the scope of analytes.

Exploring RS spectral quantitative methods with charac-teristic of high sensitivity, selectivity, simplicity, speediness,

and online is the further direction of the efforts. Firstly,prepare stable and highly active nanosol. Secondly, explorenew nanoalloy catalytic reaction including homogeneousand heterogeneous nanocatalytic reaction. Thirdly, combineDNAzyme cleaved reaction and aptamer reaction with nano-catalytic reaction to develop new nanocatalytic RS spectraltechnology.

Acknowledgments This work was supported by the National NaturalScience Foundation of China (nos. 21267004, 21165005, and 21075023),and Natural Science Foundation of Guangxi (2013GXNSFGA070003).

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